Monoclonal antibody against chicken type IX collagen: preparation, characterization, and recognition of the intact form of type IX collagen secreted by chondrocytes.

A series of monoclonal antibodies was prepared against the pepsin-resistant fragment of type IX collagen designated HMW. One of these antibodies (called 2C2) was selected for further analysis. Antibody 2C2 showed no cross-reactivity with other collagen types by inhibition enzyme-linked immunosorbent assays. It recognized an epitope present in native HMW, but failed to recognize any of the three chains of HMW fractionated after denaturation followed by reduction and alkylation of interchain disulfide bridges. Electron microscopic observations after rotary shadowing showed that the location of the epitope for antibody 2C2 was close to the carboxy-terminus of HMW. Immunofluorescent staining of sections of embryonic and adult cartilage with antibody 2C2 after removal of proteoglycans by testicular hyaluronidase digestion showed that type IX collagen is distributed throughout the cartilage matrix, and is not present in other connective tissues or skeletal muscle. The intact type IX collagen molecule, which was secreted by a suspension culture of freshly isolated embryonic chick chondrocytes, was recognized by rotary shadowing in the presence of antibody 2C2 after first precipitating the procollagens from the culture medium with ammonium sulfate (30%). Two different collagenous molecules were present in the precipitate: a longer molecule of type II procollagen (average length, 335 nm) with both amino- and carboxy-propeptides still remaining uncleaved, and a shorter molecule (average length, 190 nm) which was identified as type IX collagen. Antibody 2C2 consistently bound to the shorter molecules at a site located 136 nm from a distinctive knob at one end of the molecule, and did not bind to any specific site on the type II procollagen molecules. The structure of the intact type IX collagen molecule with the location of both collagenous and noncollagenous domains was as predicted after converting the nucleotide sequence of a cDNA clone encoding for one of the chains of type IX collagen to an amino acid sequence (Ninomiya, Y., and B. R. Olsen, 1984, Proc. Natl. Acad. Sci. USA, 81:3014-3018).

Malate content of picoliter samples of Raphanus sativus cytoplasm.

Malate, which plays many essential roles in plant metabolism, is a potent in vitro inhibitor of the cytosolic enzyme phosphoenolpyruvate carboxylase (PEPC). Because PEPC activity leads to malate biosynthesis, malate is assumed to attenuate its own synthesis in situ. To test this hypothesis, we measured directly the malate content of picoliter samples of Raphanus root-hair cytoplasm using quantitative histochemical techniques. We also obtained an estimate for malate accumulation in these cells. These values were compared with the PEPC activity of individual root hairs (less than 2 ng). The results indicate that high cytoplasmic malate concentration does not severely inhibit PEPC in situ. We suggest that the focus for studies on the regulation of organic anion accumulation be on the interactive effects of malate and other PEPC effectors.

Soft tissue fibrosarcoma. A case report and review of the literature.

The soft tissue fibrosarcoma usually presents as an enlarging, painless mass. Pain is usually a result of pressure on surrounding structures. Fibrosarcomas arise from connective tissue and demonstrate no calcification on x-ray studies. These expansile tumors are firm, round or lobulated, and well encapsulated. Their level of malignancy is graded on the basis of various degrees of differentiation of the anaplastic spindle-cells of which they are composed. Treatment generally consists of wide excision or radical local resection. Five-year survival estimates vary from 60 to 90 percent. The lung is the usual site of metastasis. A case of subungual fibrosarcoma of the great toe is presented.

Adherence and colonization of Mycoplasma pulmonis to genital epithelium and spermatozoa in rats.

Mycoplasma pulmonis is the etiological agent of a naturally occurring genital disease in rats. Transmission and scanning electron microscopic evaluations of the genital tracts of naturally and experimentally infected female rats show M. pulmonis in close association with both squamous and nonsquamous epithelial cells, although more frequently with the latter. In contrast to other species of mycoplasmas, M. pulmonis adhesion to epithelial cells appears to be mediated by a generalized interaction of the mycoplasma membrane with the host cell membrane, rather than by a specialized attachment tip. Extensive studies of all levels of the male genital tract have not yet been performed, but M. pulmonis can be demonstrated in the urethra and epididymis in animals showing evidence of chronic inflammation. Adherence of M. pulmonis to rat spermatozoa in vitro is associated with a decrease in motility. Addition of anti-M. pulmonis antibody following organism attachment results in marked agglutination of the spermatozoa. Further study of mechanisms involved in M. pulmonis adherence and subsequent mycoplasma host cell interactions is expected to contribute to an understanding of mechanisms involved in reproductive failure.